Multivalent Molecular Tweezers Disrupt the Essential NDC80 Interaction with Microtubules.

Binding of microtubule filaments by the conserved Ndc80 protein is required for kinetochore-microtubule attachments in cells and the successful distribution of the genetic material during cell division. The reversible inhibition of microtubule binding is an important aspect of the physiological error correction process. Small molecule inhibitors of protein-protein interactions involving Ndc80 are therefore highly desirable, both for mechanistic studies of chromosome segregation and also for their potential therapeutic value. Here, we report on a novel strategy to develop rationally designed inhibitors of the Ndc80 Calponin-homology domain using Supramolecular Chemistry. With a multiple-click approach, lysine-specific molecular tweezers were assembled to form covalently fused dimers to pentamers with a different overall size and preorganization/stiffness. We identified two dimers and a trimer as efficient Ndc80 CH-domain binders and have shown that they disrupt the interaction between Ndc80 and microtubules at low micromolar concentrations without affecting microtubule dynamics. NMR spectroscopy allowed us to identify the biologically important lysine residues 160 and 204 as preferred tweezer interaction sites. Enhanced sampling molecular dynamics simulations provided a rationale for the binding mode of multivalent tweezers and the role of pre-organization and secondary interactions in targeting multiple lysine residues across a protein surface.

Aromatic amines leachate from cigarette butts into aquatic environments: Is there risk for water organisms?

There are many toxics, such as aromatic amines (AAs), in cigarette butts (CBs). As CBs are the most abundant litter worldwide, these chemicals may leach into water bodies. In the present work, for the first time, the levels of AAs leachates from CBs in distilled water (DW) and river water (RW) samples were evaluated at different exposure times ranging from 15 min to 30 days. The mean leachate levels of AAs in DW and RW samples were in the range of 0.2-566 and 0.2-596 ng L-1, respectively, with overall mean values of 569 and 556 ng L-1. There was no significant difference (p > 0.05) between the total AAs levels as well as the level of each examined AA in DW and RW samples. Aniline (ANL) had the highest leaching rate from CBs into water. The mean leachates of AAs from CBs into water were ranked as: ANL> 1-naphthylamine (1-NA)> 2-naphthylamine (2-NA) > 2,6-dimethylaniline (2, 6-DMA)> ∑toluidine (∑TOL)> o-anisidine (o-ASD)> ∑aminobiphenyl (∑ABP). Ecological risk assessment showed that ∑7AAs, ANL, p-TOL, o-TOL, 2-NA, and ∑ABP had medium risks to sensitive crustaceans and fish. As AAs are not the only hazardous chemicals which may leach from CBs into aquatic environments, restrictions on littering CBs into the environment are required due to the release of different toxics ultimately causing adverse effects on aquatic organisms.

Making Every Single Puff Count-Simple and Sensitive E-Cigarette Aerosol Sampling for GCxIMS and GC-MS Analysis.

The analysis of the aerosol from tobaccoless electronic cigarettes (e-cigarettes) is an important part of understanding their impact on human health, yet sampling aerosol from e-cigarettes is still considered a challenge. It lacks a standard method for research and quality control and there are a variety of methods. However, few are simple and inexpensive, and none have been suggested for the use with gas chromatography coupled ion mobility spectrometry (GCxIMS). This work presents and evaluates such a setup made from standard lab equipment to quickly collect a quantitative sample from the aerosol of a single puff (5 s totaling 125 mL). The aerosol condensates directly in the cooled headspace (HS) vial, which is analyzed in the HS-GCxIMS or mass spectrometer (HS-GC-MS). The combined use of GC-MS and GCxIMS allows the simple and sensitive identification of unknown substances in complex mixtures and the identification of degradation products in the aerosols. A calibration of 26 flavor compounds (0.2-20 µg/g) was created using single puffs of a spiked, flavorless commercial refill solution and 2-alkanones as internal standards. This sensitive but easily reproducible setup enables a wide range of further investigations, even for labs that were previously unable to afford it.

Quantitation of Flavor Compounds in Refill Solutions for Electronic Cigarettes Using HS-GCxIMS and Internal Standards.

New regulations on the use of flavor compounds in tobaccoless electronic cigarettes require comprehensive analyses. Gas chromatography coupled ion mobility spectrometry is on the rise as an analytical technique for analyzing volatile organic compounds as it combines sensitivity, selectivity, and easy usage with a full-range screening. A current challenge is the quantitative GCxIMS-analysis. Non-linear calibration methods are predominantly used. This work presents a new calibration method using linearization and its corresponding fit based on the relation between the reactant and analyte ions from the chemical ionization. The analysis of e-liquids is used to compare the presented calibration with an established method based on a non-linear Boltzmann fit. Since e-liquids contain matrix compounds that have been shown to influence the analyte signals, the use of internal standards is introduced to reduce these effects in GCxIMS-analysis directly. Different matrix mixtures were evaluated in the matrix-matched calibration to improve the quantitation further. The system's detection and quantitation limits were determined using a separate linear calibration. A matrix-matched calibration series of 29 volatile compounds with 12 levels were used to determine the concentration of these substances in a spiked, flavorless e-liquid and a banana-flavored e-liquid, validating the quality of the different calibrations.

Analysis of volatile metabolites from in vitro biofilms of Pseudomonas aeruginosa with thin-film microextraction by thermal desorption gas chromatography-mass spectrometry.

Cystic fibrosis (CF) is an autosomal recessive inherited disease which leads to a production of thickened mucus in the airways. These conditions are conducive to poly-microbial infections, like chronic lung infection, in which Pseudomonas aeruginosa (P. aeruginosa) is the major pathogenic bacterium colonizing CF lungs at the end of the lifetime of CF patients. This in vitro study uses a P. aeruginosa biofilm model under partly cystic fibrosis conditions, with a sampling of volatile extracellular metabolites. The gas sampling was done with thin-film microextraction (TFME) and commercial polydimethylsiloxane (PDMS) films, whereas the analysis of loaded films was done by gas chromatography coupled to quadrupole mass spectrometry and thermodesorption (TD-GC-qMS). For this purpose, two commercially available films were characterized by means of thermogravimetry coupled to a qMS with atmospheric pressure photo ionization (TG-APPI-qMS), regarding homogeneity and temperature stability. The selected film was cleaned using a method developed in this study. The TD-GC-qMS method was successfully used for standards of volatile metabolites which were known to be produced by P. aeruginosa. Limits of detection and quantification of the method for middle and less polar compounds in low nanomolar range (0.5 nM and 1.5 nM) were achieved. The developed method was finally applied to investigate the extracellular volatile metabolites produced by biofilms of the strain P. aeruginosa DSM 50071 under aerobic and anaerobic conditions. In sum, eleven metabolites could be found under both conditions. Furthermore, it was shown in this study that different oxygen conditions (aerobic and anaerobic) resulted in emitting different extracellular volatile metabolites. Specific metabolites, like 1-undecene (aerobic) and 2-undecanone (anaerobic), could be identified. The results are promising, in that the biofilm model may be applicable for the identification of P. aeruginosa under clinical conditions. Furthermore, the model could be the basis for studying extracellular volatile metabolites from different mono- or co-cultures of various bacteria, as well as the implementation of pulmonary conditions, like these in CF lungs. This possibility allows the development of a non-invasive "at-bedside" breath analysis method for CF patients in focus of various bacterial infections. Graphical abstract.

Detecting Early Markers of Ventilator-Associated Pneumonia by Analysis of Exhaled Gas.

OBJECTIVES: The detection of microbial volatile organic compounds or host response markers in the exhaled gas could give an earlier diagnosis of ventilator-associated pneumonia. Gas chromatography-ion mobility spectrometry enables noninvasive, rapid, and sensitive analysis of exhaled gas. Using a rabbit model of ventilator-associated pneumonia we determined if gas chromatography-ion mobility spectrometry is able to detect 1) ventilator-associated pneumonia specific changes and 2) bacterial species-specific changes in the exhaled gas. DESIGN: Experimental in vivo study. SETTING: University research laboratory. SUBJECTS: Female New Zealand White rabbits. INTERVENTIONS: Animals were anesthetized and mechanically ventilated. To induce changes in the composition of exhaled gas we induced ventilator-associated pneumonia via endobronchial instillation of either Escherichia coli group (n = 11) or Pseudomonas aeruginosa group (n = 11) after 2 hours of mechanical ventilation. In a control group (n = 11) we instilled sterile lysogeny broth endobronchially. MEASUREMENTS AND MAIN RESULTS: Gas chromatography-ion mobility spectrometry gas analysis, CT scans of the lungs, and blood samples were obtained at four measurement points during the 10 hours of mechanical ventilation. The volatile organic compound patterns in the exhaled gas were compared and correlated with ventilator-associated pneumonia severity. Sixty-seven peak areas showed changes in signal intensity in the serial gas analyses. The signal intensity changes in 10 peak regions differed between the groups. Five peak areas (P_648_36, indole, P_714_278, P_700_549, and P_727_557) showed statistically significant changes of signal intensity. CONCLUSIONS: This is the first in vivo study that shows the potential of gas chromatography-ion mobility spectrometry for early detection of ventilator-associated pneumonia specific volatile organic compounds and species differentiation by noninvasive analyses of exhaled gas.

Time-of-flight ion mobility spectrometry in combination with laser-induced fluorescence detection system.

A laser-induced fluorescence (LIF) was used as a complimentary detection system for time-of-flight ion mobility spectrometry (TOF-IMS). A LIF detection system is potentially faster than a conventional electrometer detector and can provide additional (to usual for IMS drift time) analytical information, namely wavelength of fluorescence maxima and fluorescence lifetime. Therefore, better discrimination ability can be expected. Additionally, the combination of IMS and LIF operates at atmospheric pressure. This allows fluorescence measurements of specified ions and ion clusters, which would not survive in a mass spectrometer. An IMS drift cell of open design with both the electrometer and LIF detectors was designed. The feasibility of IMS-LIF was demonstrated on the example of the Xanthene dye Rhodamine 6G (R6G). Electrospray was used as an ionization source. The release and desolvation of R6G ions from the electrospray with following IMS-LIF analysis were demonstrated. The effects of experimental parameters (e.g., ion gate and drift voltages, distance to ESI emitter) are demonstrated and discussed. The obtained results are promising enough to ensure the potential of LIF as a complimentary/alternative detection system for time-of-flight ion mobility spectrometry.

Identification of terpenes and essential oils by means of static headspace gas chromatography-ion mobility spectrometry.

Static headspace gas chromatography-ion mobility spectrometry (SHS GC-IMS) is a relatively new analytical technique that has considerable potential for analysis of volatile organic compounds (VOCs). In this study, SHS GC-IMS was used for the identification of the major terpene components of various essential oils (EOs). Based on the data obtained from 25 terpene standards and 50 EOs, a database for fingerprint identification of characteristic terpenes and EOs was generated utilizing SHS GC-IMS for authenticity testing of fragrances in foods, cosmetics, and personal care products. This database contains specific normalized IMS drift times and GC retention indices for 50 terpene components of EOs. Initially, the SHS GC-IMS parameters, e.g., drift gas and carrier gas flow rates, drift tube, and column temperatures, were evaluated to determine suitable operating conditions for terpene separation and identification. Gas chromatography-mass spectrometry (GC-MS) was used as a reference method for the identification of terpenes in EOs. The fingerprint pattern based on the normalized IMS drift times and retention indices of 50 terpenes is presented for 50 EOs. The applicability of the method was proven on examples of ten commercially available food, cosmetic, and personal care product samples. The results confirm the suitability of SHS GC-IMS as a powerful analytical technique for direct identification of terpene components in solid and liquid samples without any pretreatment. Graphical abstract Fingerprint pattern identification of terpenes and essential oils using static headspace gas chromatography-ion mobility spectrometry.

Fast detection of coliform bacteria by means of gas chromatography-differential mobility spectrometry.

In this study, we demonstrate that the combination of an enzymatic method (based on Colilert-18 medium) and gas chromatography-differential mobility spectrometry (GC-DMS) can reduce the time required for detection of coliform bacteria (including Escherichia coli) from 18 to 2.5 h. The presented method includes the incubation (~2.5 h) of the sample containing coliform bacteria in Colilert-18 medium. The incubation time of 2.5 h is required for the activation of the ß-galactosidase enzyme. Produced during the incubation biomarker o-nitrophenol (ONP) can be detected by means of GC-DMS within just 200 s. The detection limit for ONP was 45 ng (on-column). The method developed in this work provides significantly shorter analysis time compared with standard methods, and can be potentially adapted to the field conditions. Therefore, this method is a promising tool for an early detection of coliform bacteria (including E. coli). Graphical Abstract Fast detection of coliform bacteria by means of GC-DMS.

Low-temperature plasma ionization differential ion mobility spectrometry.

A low-temperature plasma (LTP) was used as an ionization source for differential ion mobility spectrometry (DMS) for the first time. This ionization source enhances the potential of DMS as a miniaturized system for on-site rapid monitoring. The effects of experimental parameters (e.g., discharge/carrier gas composition and flow rate, applied voltage) on the analysis of model aromatic compounds were investigated and discussed. It was found that the nature of reactant ion positive (RIP) is dependent on the discharge/carrier gas composition. The best response to the analyte was achieved when pure nitrogen was used as the discharge/carrier gas. The ability to perform analysis with zero helium consumption is especially attractive in view of the potential application of LTP-DMS for online (and on-site) monitoring. Analytical performance was determined with six environmentally relevant model compounds (benzene, toluene, ethylbenzene, p-xylene, 1,2,4-trimethylbenzene, and naphthalene) using LTP and directly compared to APPI and APCI ((63)Ni) ionization sources. When LTP was coupled to DMS, calculated LOD values were found to be in the range of 35-257 ng L(-1) (concentration in the carrier gas). These values are competitive with those calculated for two DMS equipped with traditional ionization sources (APPI, (63)Ni). The obtained results are promising enough to ensure the potential of LTP as ionization source for DMS.

Advanced remediation in the presence of ferrous iron and carbonate-containing water by oxygen-induced oxidation of organic contaminants.

Mechanistic investigations of an environmentally friendly and easy-to-implement oxidation method in the remediation of contaminated anoxic waters, i.e. groundwater, through the sole use of oxygen for the oxygen-induced oxidation of pollutants were the focus of this work. This was achieved by the addition of O2 under anoxic conditions in the presence of ferrous iron which initiated the ferrous oxidation and the simultaneous formation of reactive •OH radicals. The involvement of inorganic ligands such as carbonates in the activation of oxygen as part of the oxidation of Fe2+ in water was investigated, too. The formation of •OH radicals, was confirmed in two different, indirect approaches by a fluorescence-based method involving coumarin as •OH scavenger and by the determination of the oxidation products of different aromatic VOCs. In the latter case, the oxidation products of several typical aromatic groundwater contaminants such as BTEX (benzene, toluene, ethylbenzene, xylenes), indane and ibuprofen, were determined. The influence of other ligands in the absence of bicarbonate and the effect of pH were also addressed. The possibility of activation of O2 in carbonate-rich water i.e. groundwater, may also potentially contribute to oxidation of groundwater contaminants and support other primary remediation techniques.

Elucidating nicotine transfer into water environments via cigarette butt remaining parts.

Nicotine, the main alkaloid in tobacco, enters water environments through discarded cigarette butts (CBs), possibly causing detrimental effects. However, there is no comprehensive investigation on the long-term leaching of nicotine from the different CBs parts. Therefore, in the present study, the ecological risk and the leachate levels of nicotine from different CBs parts were investigated. Freshly smoked CBs, aged CBs collected from streets, remaining tobacco and ash of freshly smoked CBs, and filter plus paper of freshly smoked CBs were evaluated for the leachate experiments. The order of nicotine leachate from different types of CBs and parts investigated were as remaining tobacco plus ash of freshly smoked CBs > freshly smoked CBs > aged CBs > filter plus paper of freshly smoked CBs with the ranges of 5.73-17.34, 0.36-8.6, 0.31-4.12, and 0.17-2.79 mg of nicotine per g of CB or remaining parts (mg g-1), respectively. The ecological risk assessment revealed that nicotine leachates from all the CBs types or their remaining parts could be highly hazardous to fish, cladocerans, algae, and Daphnia magna. Based on the mean leachate levels of nicotine via freshly smoked CBs at exposure times of 1 min to 1 month and the estimated number of littered CBs every year on a worldwide scale, freshly smoked CBs may release 380-7065 tons of nicotine into water environments.

A novel coupling technique based on thermal desorption gas chromatography with mass spectrometry and ion mobility spectrometry for breath analysis.

Exhaled breath analysis is evolving into an increasingly important non-invasive diagnostic tool. Volatile organic compounds (VOCs) in breath contain information about health status and are promising biomarkers for several diseases, including respiratory infections caused by bacteria. To monitor the composition of VOCs in breath or the emission of VOCs from bacteria, sensitive analytical techniques are required. Next to mass spectrometry, ion mobility spectrometry (IMS) is considered a promising analytical tool for detecting gaseous analytes in the parts per billion by volume to parts per trillion by volume range. This work presents a new, dual coupling of thermal desorption gas chromatography to a quadrupole mass spectrometer (MS) and an IMS by operating a simple splitter. Nearly identical retention times can be reached in the range of up to 30 min with slight deviations of 0.06 min-0.24 min. This enables the identification of unknown compounds in the IMS chromatogram using unambiguous mass spectral identification, as there are still no commercially available databases for IMS. It is also possible to discriminate one of the detectors using the splitter to improve detection limits. Using a test liquid mixture of seven ketones, namely 2-butanone, 2-pentanone, 2-hexanone, 2-heptanone, 2-octanone, 2-nonanone, and 2-decanone with a concentration of 0.01 g l-1reproducibilities ranging from 3.0% to 7.6% for MS and 2.2%-5.3%, for IMS were obtained, respectively. In order to test the system optimized here for the field of breath analysis, characteristic VOCs such as ethanol, isoprene, acetone, 2-propanol, and 1-propanol were successfully identified in exhaled air using the dual detector system due to the match of the corresponding IMS, and MS spectra. The presented results may be considered to be a starting point for the greater use of IMS in combination with MS within the medical field.

Atmospheric pressure photoionization - High-field asymmetric ion mobility spectrometry (APPI-FAIMS) studies for on-site monitoring of aromatic volatile organic compounds (VOCs) in groundwater.

A continuously operating system for monitoring groundwater contamination by aromatic VOCs has been developed. For this purpose, a novel gas-water separation unit was to be used in combination with APPI-FAIMS. The gas-water separation unit successfully reduced the humidity in the sample flow to ≤1.6 ppmv prior to analyte ionization. Initially, toluene was selected as a model aromatic VOC. The quantitative response of toluene, as a single VOC in water (LOD <1 mg L-1), was used to investigate the feasibility of the monitoring system and the effect of humidity on the signal produced by the APPI-FAIMS. With humidity increase (up to 400 ppmv) an increase of the toluene signal for about 30% was observed, including the possible formation and detection of water clusters and toluene-water clusters. Similar effects were noted in the case of benzene. However, for the detection of single contaminants such as indane and trimethylbenzenes (TMBs) this was not observed even at relative high humidity (500 ppmv). Additionally, on-site, continuous, groundwater monitoring of the aromatic VOCs contamination was carried out successfully with the gas-water separation APPI-FAIMS at low humidity (0.3-1.6 ppmv) allowing simplified monitoring of a specific, total aromatic VOCs signal in groundwater.

Characterization of the Extracellular Volatile Metabolome of Pseudomonas Aeruginosa Applying an in vitro Biofilm Model under Cystic Fibrosis-Like Conditions.

BACKGROUND: Cystic fibrosis (CF) is an autosomal recessive hereditary disease that leads to the production of thickened mucus in the lungs, favouring polymicrobial infections, such as chronic lung infections with the bacterial opportunistic pathogen Pseudomonas aeruginosa. METHOD: A biofilm model in combination with an adapted sampling and GC-MS analysis method were applied to in vitro studies on different variables influencing the composition of the extracellular volatile metabolome of P. aeruginosa. RESULTS: A significant influence on the metabolome could be demonstrated for the culture medium as well as the atmosphere during cultivation (aerobic or anaerobic). Furthermore, a significant influence of the mucoid (alginate-overproducing) phenotype of the bacterium on quantity and composition of volatile organic compounds could be observed. Based on the results a solid culture medium was developed to simulate the nutrient conditions in the lungs of a CF patient. The extracellular volatile metabolome of bacterial strains P. aeruginosa ATCC 10145, PAO1 and FRD1 was characterized under CF-like conditions. CONCLUSIONS: Bacterial strain-dependent metabolites were identified. When P. aeruginosa PAO1 and FRD1 clinical isolates were compared, 36 metabolites showed significant variations in intensities. When the clinical isolates were compared with the reference strain (P. aeruginosa ATCC 10145), 28 metabolites (P. aeruginosa PAO1) and 70 metabolites (P. aeruginosa FRD1) were determined whose peaks showed significant deviation (p > 95%) in intensity. Furthermore, the bacterial strains could be differentiated from each other by means of two principal components.

Strategy for the identification of flavor compounds in e-liquids by correlating the analysis of GCxIMS and GC-MS.

This work presents a strategy to correlate the results from gas chromatography coupled ion mobility spectrometry (GCxIMS) and mass spectrometry (GC-MS) to enable a simpler and cheaper analysis of flavor compounds in e-liquids. The use of the retention index for GCxIMS measurements was validated for its application to correlate results with GC-MS data. The easy detection of the GCxIMS for substances at concentrations as low as 1 µg/L can therefore be combined with the identification power of the MS. The use of the MS' mass signals and wide-spread availability of mass spectra libraries reduces the effort necessary to choose the correct reference standards for the identification of unknown substances. Between both detectors, correlating of the retention time indices was achieved for ± 1%. 2-Alkanones were used as an alternative reference point for the IMS and the well-established alkanes for the MS. The application on flavor compounds in e-liquids shows equal or better results than those presented for more complex, hardware-based correlations like line splitting. Additionally, the inverted reduced mobility combined with the retention index of a non-polar column enables simple extrapolation for the confirmation of expected substances as well as the use in a transferable database. For the first time, this comprehensive application allows an extensive, simplified, and cheap identification of flavor compounds in e-liquids by GCxIMS and GC-MS.

Polycyclic aromatic hydrocarbons (PAHs) leachates from cigarette butts into water.

Cigarette butts (CBs) are the most common littered items in the environment and may contain high amounts of polycyclic aromatic hydrocarbons (PAHs) from incomplete tobacco leave burning. The potential relevance of PAHs stemming from CBs for aquatic systems remain unclear since to date there is no systematic study on PAHs leaching from CBs. Therefore, in this study the leaching concentrations of 16 EPA-PAHs (except benzo(ghi)perylene) in 3 different types of water were measured. The concentrations of ΣPAHs leachates from 4 h to 21 days ranged from 3.9 to 5.7, 3.3-5.5, and 3.0-5.0 µg L-1 for deionized, tap, and river waters, respectively. For all contact times, there were no substantial differences of the leachate concentrations of PAHs among different water types. Lighter PAHs had the highest concentrations among the detected PAHs and they were detected in the leachates already after 4 h. Concentrations of indeno(1,2,3-cd)pyrene, and dibenz(a,h)anthracene were below the limit of detection in all water samples at different contact times. At all contact times naphthalene and fluorene had the highest concentrations among the studied PAHs. Tap and river water samples with addition of sodium azide as chemical preservative contained significantly higher concentration of ΣPAHs. Our leaching data showed that leached concentrations of PAHs exceeded the Water Framework Directive (WFD) standards and considering the number of CBs annually littered this may pose a risk to aquatic organisms and potentially also humans.

Analysis of microbial extracellular polysaccharides in biofilms by HPLC. Part I: Development of the analytical method using two complementary stationary phases.

The investigation of microbial extracellular polymeric substances (EPS) is helpful for the implementation of analytical methods which are suitable for biofilm analysis in order to understand the architecture and function of biofilms. A procedure for the qualitative and quantitative determination of various monosaccharides, oligosaccharides and uronic acids as important components of the carbohydrate fraction of microbial EPS by high-performance liquid chromatography (HPLC) and refractive index (RI)/UV detection is presented. Porous graphitic carbon and lead-form cation-exchanger have been examined as stationary phases. Therefore, two complementary HPLC methods are presented. To simulate the conditions of hydrolysis, the influences of various salts, acids and alkalis as matrix components have been investigated. Furthermore, the dependencies on the pH value and temperature of the mobile phase have been thoroughly studied. The results showed that the lead-form cation-exchanger is suitable for the separation of the neutral monosaccharides. However, for direct analysis after acidic hydrolysis with H(2)SO(4), HCl or trifluoroacetic acid, an additional purification step, e.g., precipitation or lyophilization, is necessary when the cation-exchanger is used. With the exception of hydrolysis with HCl, the porous graphitic carbon stationary phase can be used without any further purification step and is appropriate for the separation of uronic acids and their gamma-lactones. Additionally, the separation of a single monosaccharide and its derivatives is possible. Analytical parameters including the sensitivities, repeatabilities, limits of detection and limits of quantification of both HPLC methods using the RI detector are presented. The optimized method has been applied for the characterization of alginates and is also suitable for other extracellular polysaccharides in biofilms.

Direct inlet probe ion mobility spectrometry.

Direct inlet probe (DIP) was used as an introduction and a pre-separation step for atmospheric pressure photoionization time-of-flight ion mobility spectrometry (APPI-TOF-IMS) for the first time. IMS is an analytical technique used to separate and identify ionized molecules in the gas phase and under atmospheric pressure based on their mobility. The utilization of DIP prior to IMS gives the possibility to introduce the analytes into the gas phase and provides an additional separation based on their vapor pressure. The proof-of-principle study was done on example of eight polycyclic aromatic hydrocarbons (PAHs) with the ring number from 2 to 5, namely naphthalene, fluorene, anthracene, phenanthrene, pyrene, fluoranthene, benzo[a]pyrene, and benzo[k]fluoranthene. All these compounds are included in EPA priority pollutant list. Moreover, benzo[a]pyrene and benzo[k]fluoranthene are marked by EPA as probably carcinogen compounds and also included into SCF and EU lists. To increase the sensitivity of DIP-APPI-IMS the analysis was performed using a dopant assisted ionization method (benzene, 74mgL-1 in N2). It was found that the heating rate of the interface plays a crucial role for the whole analytical procedure. To prove the ability of this method to analyze PAHs in the mixture, the mixtures containing up to five PAHs were analyzed. The LODs for the analyzed compounds obtained with DIP-APPI-IMS were found to be in the tens- or hundreds-of-microgram-per-liter range. The obtained results are promising enough to ensure the potential of DIP as an introduction and a pre-separation step for ion mobility based methods.

X-ray ionization differential ion mobility spectrometry.

X-ray was utilized as an ionization source for differential ion mobility spectrometry (DMS) for the first time. The utilization of this ionization source increases the potential of DMS system for on-site based applications. The influence of experimental parameters (e.g. accelerating voltage, filament current, and separation field) on the analysis of model compounds was investigated and discussed. It was found that both the positive and the negative reactive ion peaks [RIP(+) and RIP(-)] formed during X-ray ionization are identical with those observed with the traditional 63Ni radioactive ion source. This is especially notable for RIP(-), because the chemistry provided by other nonradioactive sources in the negative mode is more complicated or even different than that observed with a 63Ni source. Increase of either filament current or accelerating voltage resulted in increased intensity of both RIP(+) and RIP(-). However, because of the materials used for construction of X-ray adapter the maximal level of filament current and accelerating voltage used in this study were limited to 700mA and 5kV, respectively. Analytical performance was determined with two model compounds (acetone and methyl salicylate) using X-ray and directly compared to 63Ni ionization source. When X-ray was coupled to DMS, calculated LOD values were found to be within the range of 0.17-1.52ppbv/v (concentration in the carrier gas). These values are competitive with those calculated for DMS equipped with traditional 63Ni radioactive ionization source. The obtained results are promising enough to ensure the potential of X-ray as ionization source for DMS.